WO2007084818A2 - Compositions pharmaceutiques et méthodes permettant d'obtenir et de maintenir un taux de cuivre cible, stable, et de prévenir et de traiter des maladies du système nerveux central liées au taux de cuivre sérique - Google Patents

Compositions pharmaceutiques et méthodes permettant d'obtenir et de maintenir un taux de cuivre cible, stable, et de prévenir et de traiter des maladies du système nerveux central liées au taux de cuivre sérique Download PDF

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WO2007084818A2
WO2007084818A2 PCT/US2007/060345 US2007060345W WO2007084818A2 WO 2007084818 A2 WO2007084818 A2 WO 2007084818A2 US 2007060345 W US2007060345 W US 2007060345W WO 2007084818 A2 WO2007084818 A2 WO 2007084818A2
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copper
zinc
disease
formulation
composition
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PCT/US2007/060345
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WO2007084818A3 (fr
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Steve H. Kanzer
George J. Brewer
Nicholas Stergis
John S. Althaus
Charles S. Bisgaier
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Pipex, Inc.
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Priority to AU2007205996A priority Critical patent/AU2007205996A1/en
Priority to CA002675230A priority patent/CA2675230A1/fr
Priority to EP07718298A priority patent/EP1993607A4/fr
Priority to JP2008550493A priority patent/JP2009523175A/ja
Publication of WO2007084818A2 publication Critical patent/WO2007084818A2/fr
Publication of WO2007084818A3 publication Critical patent/WO2007084818A3/fr

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Definitions

  • the present invention relates to pharmaceutical products and methods for treating excessive metal buildup or metal malabsorption in animals and humans.
  • the invention has particular applicability to treatment of Wilson's Disease in humans and will be described in connection with such utility, although other utilities are contemplated including treatment of other neurological diseases caused by excessive copper accumulation in the brain and other organs and/or intraday fluctuations in levels of free copper in the serum and central nervous system (CNS), including but not limited to Alzheimer's Disease, Parkinson's Disease, amyotrophic lateral sclerosis (ALS) or Lou Gehrig's Disease, dementia, Huntington's Disease, and schizophrenia, as well as neuromuscular diseases associated with abnormal accumulation of copper associated proteins in the body, such as juvenile and sporadic inclusion body myositis and myositis of the elderly, and cardiovascular diseases such as atherosclerosis, stroke, and peripheral vascular disease.
  • CNS central nervous system
  • the invention also may be used for the treatment of neurological and psychiatric manifestations of hepatic diseases associated with impaired liver copper excretion, such as colangitis, hepatitis and cirrhosis, for example, in which free or loosely bond serum or CSF copper is elevated.
  • Copper is a trace element that is essential to life. Despite its essentiality, however, copper also is an extremely reactive oxidative species that has the potential to be very toxic to cells, proteins, and organ systems such as the liver, brain and vasculature.
  • cuproproteins include, but are not limited to, matrix metalloprotein, ceruloplasmin, copper/zinc superoxide dismutase, amyloid precursor protein, apolipoprotein E, tau, homocysteine, albumin and chaperone for copper zinc superoxide dismutase, to name a few.
  • matrix metalloprotein ceruloplasmin
  • copper/zinc superoxide dismutase copper/zinc superoxide dismutase
  • amyloid precursor protein apolipoprotein E
  • tau amyloid precursor protein
  • homocysteine albumin
  • chaperone for copper zinc superoxide dismutase to name a few.
  • One of the most problematic and potentially toxic sources of copper for humans is the abundance of toxic copper ions that exists in drinking water systems.
  • the copper in drinking water occurs in the form of cupric ion (Cu +2 ) in either an unbound form or in a form complexed only loosely with organic ligands.
  • Copper ions are generally more bioavailable in water than they are in food; there may be components in food that can influence the metabolism, absorption and mobilization of copper in human diets. Absorption of Copper by the Human Body In humans, dietary copper is absorbed from the stomach and small intestine.
  • Ceruloplasmin is a cysteine-rich glycoprotein with many free sulfhydryl groups that serve as binding points for metals. Ceruloplasmin can bind copper or zinc, but has a stronger affinity for copper (Cousins, 1985). Ceruloplasmin is synthesized on membrane-bound polyribosomes of liver parenchymal cells and secreted into the plasma. Copper that enters the portal circulation from the intestine is transported directly to the liver. Copper released from the liver is transported in the bloodstream to other organs, including the kidney and brain. The synthesis of ceruloplasmin is controlled by interleukin-I via glucagon or glucocorticoid (Cousins, 1985).
  • Circulating copper levels are elevated in pregnant women because hormonal changes associated with pregnancy stimulate ceruloplasmin synthesis (Solomons, 1985). Ceruloplasmin levels may be useful as an indicator of copper status (Mendez et al., 2004). Recently, several copper transporters involved in copper uptake and transport by cells have been identified (Bauerly et al., 2005). Copper transporter- 1 (Crtl) is a copper import protein that is copper-specific, and is believed to mediate copper uptake into the small intestine (Lee et al., 2002). Crtl is expressed in the enterocytes of the small intestine and in enterocyte-like Caco-2 cells in culture (Klomp et al., 2002; Kuo et al., 2001).
  • ATP7A The copper efflux protein, ATP7A, is thought to mediate copper efflux across the plasma membrane during copper excess in transfected cells (Petris et al., 1996). Menkes disease, characterized by excessive copper accumulation in the intestine and systemic copper deficiency, is a consequence of a defect in ATP7A (Schaefer and Gitlin, 1999). ATP7B, with functional similarity to ATP7A, exports copper into bile for excretion (Roelofsen et al., 2000). ATP7B is localized primarily in the liver with lower expression found in the intestine, kidney and placenta (Lockhart et al., 2000).
  • a defect in ATP7B results in Wilson's disease, characterized by copper toxicity (due to liver copper accumulation as a result of impaired biliary copper excretion) and liver damage.
  • Metabolism and Excretion of Copper The liver and intestine play key roles in copper metabolism. Copper is taken up by hepatocytes from the portal circulation. Inside the hepatocytes, copper is bound to metallothionein, a protein that also binds zinc, iron and mercury. Copper can be released from hepatocytes into the general circulation to be transported to other tissues, or it can be excreted from the liver in bile (Cousins, 1985). The major route of excretion is in the bile.
  • Nervous system effects including dementia, have been observed in individuals with copper deficiency or excess (Solomons, 1985; Harris, 1997). Effects on catecholamine metabolism likewise are involved in the nervous system abnormalities. Other physiological functions that involve copper include leukopoiesis, skeletal mineralization, connective tissue synthesis, melanin synthesis, oxidative phosphorylation, thermal regulation, antioxidant protection, cholesterol metabolism, immune and cardiac function, and regulation of glucose metabolism. Since all of these physiological processes involve copper, any of them can be affected by the availability of copper in the body or in specific tissues. In general, deleterious effects may occur in any of these physiological processes due to either deficiency or excess of copper in the systems affected (Solomons, 1985; Harris, 1997).
  • Table 1 shows the Dietary Reference Intake (DRI) values for copper for various age groups, broken down into Estimated Average Requirements (EAR), Recommended Dietary Allowances (RDA), and Tolerable Upper Intake Levels (UL) (FNB, 2000). Values for infants were provided only as Adequate Intake values, based primarily on the content of copper in human milk.
  • the AI values are 200 ⁇ g/day for infants 0-6 months of age, and 220 ⁇ g/day for infants at 7-12 months; an estimated UL for infants could not be established (FNB, 2000).
  • Wilson's disease While the majority of persons may be able to cope with chronic exposure to toxic copper ions contained in drinking water without showing signs of disease, there are certain rare diseases in which a person's copper transport and metabolic pathways are affected by genetic mutations, such as Wilson's disease and Menkes disease.
  • the genetic mutations responsible for Wilson's disease and Menkes disease were identified for the first time in the 1990's by several groups.
  • there have been some published reports of elevated levels of serum copper in the elderly (Madaric et. al., Physiol Res, 1994; 43(2): 107- 11 and Ghayour-Mobarhan et.
  • Wilson's disease is characterized by a mutation of the gene encoding the P- type ATPase, called ATP7B.
  • Wilsons' disease patients are unable to adequately process, transport and excrete copper through the normal bile ducts of the liver.
  • copper that is newly introduced is expected to first bind to available engogenous cuproproteins having the highest affinity for copper, such as metallothionein, superoxide dismutase and albumin.
  • Free copper ions are relatively rare in serum, but copper which is "loosely bound" to various proteins and peptides can be substantial and elevated in Wilson' s disease patients and potentially also in other metabolically compromised groups such as Alzheimer's disease patients, mild cognitive impairment (MCI) patients, schizophrenia patients, dementia patients, and the elderly.
  • MCI mild cognitive impairment
  • Menkes Disease is characterized by abnormally low levels of available copper, due to the failure of intestinal cells to release copper, and results in various developmental abnormalities.
  • ceruloplasmin 650-750ug/L, 65-70%
  • albumin 120-180 ug/L, 12-18%)
  • transcuprein microglobulin
  • ferroxidase II 10 ug/L, 1%
  • extracellular SOD and histidine rich glycoproteins ⁇ 10 ug/L, ⁇ 1%
  • blood clotting factors V and VIII ⁇ 5 ug/L, ⁇ 0.5%)
  • extracellular metallothionein and anime oxidase ⁇ 1 ug/L, ⁇ 0.1%)
  • 15-6OkDa components 40 ug/L, 4%
  • small peptides and amino acids 35 ug/L, 4%
  • unbound or "free" copper ions 0.0001 ug/L, approx.
  • neurons may upregulate a variety of copper binding proteins, including APP, Amyloid beta, tau, BACEl and apoE, all of which are upregulated in Alzheimer's disease (and intracellularly in a similar fashion in the neuromuscular disease, inclusion body myositis).
  • copper binding proteins including APP, Amyloid beta, tau, BACEl and apoE, all of which are upregulated in Alzheimer's disease (and intracellularly in a similar fashion in the neuromuscular disease, inclusion body myositis).
  • Solubilized copper or copper loosely bound to small ligands such as that commonly found in tap water, is highly bioavailable (up to 65%) and, due to water fluxes in the intestines, has the capacity to overwhelm the copper homeostasis mechanisms of the gastrointestinal enterocytes and liver, and enter the portal and systemic circulation in a potentially toxic form loosely bound to albumin and other low kinetic copper binding proteins. It is an object of the present invention to provide compositions, formulations, agents and methods to protect the individual from such toxic fluxes, as further described herein. Wilson's Disease In the case of untreated Wilson's disease patients, body copper continues to accumulate, ultimately overwhelming the high affinity cuproproteins.
  • Residual copper either remains free and unbound or loosely bound to cuproproteins having low affinities to copper.
  • This pool of free, unbound or loosely bound copper is free to circulate and may cross the blood brain barrier, damaging nerve cells due to its reactivity and pro-oxidant capacity.
  • the various cuproproteins serve as a reservoir for copper and will generally be released based upon an inverse correlation with each cuproprotein's individual affinity for copper.
  • the so called "loosely bound" cuproproteins include albumin and homocysteine, for example.
  • Such cuproproteins serve as potential toxic pools of available copper as compared to high affinity copper binding proteins, such as ceruloplasmin and cu/zn superoxide dismutase.
  • Wilson's Disease Treatment for Wilson' s Disease
  • a network of cuproproteins is abnormal as a result of genetic abnormality or aging, for example, then copper in the body may not be properly bound and sequestered by cuproproteins and therefore may not be properly maintained.
  • Toxicities and oxidative capacities are most pronounced when copper is present in the body in its so-called “free", “unbound” or “loosely bound” forms.
  • Such free copper can be toxic to various organ systems, such as the liver and the brain, for example.
  • the classic example of such a disease is Wilson's disease.
  • Treatment varies for patients that are initially presenting, and they are generally treated on an acute or induction basis, with potent copper chelators and complexors, such as tetrathiomolybdate, penicillamine or trientine, each of which is intended to either remove available free copper from the body or render it unavailable for further damage.
  • potent copper chelators and complexors such as tetrathiomolybdate, penicillamine or trientine, each of which is intended to either remove available free copper from the body or render it unavailable for further damage.
  • agents commonly used for chronic or maintenance therapy include those that maintain a state of copper malabsorption, such as zinc acetate (Brewer).
  • Zinc acetate is available as a prescription and is marketed in the United States under the tradename Galzin® and in Europe under the tradename Wilzin®.
  • Zinc was used for the comprehensive treatment of Wilson's disease including initial treatment (Hoogenraad et al., Lancet, 2:1262-1263 [1978]; Hoogenraad et al., Eur. Neurol., 18:205-211 [1979]; and Hoogenraad et al., J. Neurol. ScL, 77:137-146 [1987]).
  • TM Tetrathiomolybdate
  • Such chronic maintenance therapies fail in some patients due to chronic and acute stomach and esophageal irritation and nausea commonly associated with such agents, difficulty in predicting effects and in setting an appropriate dosing regimen, and the need to continuously monitor free serum copper levels in order to assure that they are maintained within the normal range.
  • the variability of effect of such agents depends upon the timing of administration as it relates to the timing of meals, difficulties in maintaining adequate patient compliance given the daily multiple dosing regimen, possible stomach irritation and the need to time each dose at least one hour prior and three hours after meals, as well as the need to assure compliance for the entire remaining lifetime of the patient.
  • Other Disorders associated with elevated levels of free copper and/or accumulation of copper are associated with elevated levels of free copper and/or accumulation of copper.
  • Other disorders are associated with elevated levels of loosely bound, free copper, and/or accummulation of copper includes headaches, hypoglycemia, increased heart rate, nausea, anemia, hair loss, nephritis, autism, depression, hallucinations, hyperactivity, insomnia, disperception of the senses, paranoia, personality changes, psychosis, schizophrenia, mild cognitive impairment, detachment from reality, atherosclerosis, stroke, tauapathies and synucleinopathies, nonalcoholic steatohepatitis, multiple sclerosis, Alzheimer's, Parkinson's, dementia, ALS and autism which are given as exemplary.
  • Alzheimer's disease As human life span has significantly expanded over the last century, Alzheimer's disease and other neurodegenerative diseases will have a growing impact on the quality of life for a large proportion of the population. For example, Alzheimer's disease is a leading cause of dementia in the elderly, affecting 5-10% of the population over the age of 65 years. See A Guide to Understanding Alzheimer's disease and Related Disorders, edited by Jorm, New York University Press, New York (1987). Alzheimer's disease often presents with a subtle onset of memory loss followed by a slow progressive dementia over several years.
  • Alzheimer's disease has hampered the development of a model that mimics many of the changes seen in the human brain. Such a model is needed in order to identify drugs or other agents that might be useful in treating, preventing or reversing the effects of such diseases.
  • Alzheimer's disease is histopathologically characterized by the loss of particular groups of neurons and the appearance of two principal lesions within the brain, termed senile plaques and neurofibrillary tangles. See Brion et al., J. Neurochem. 60: 1372-1382 (1993). Senile plaques occur in the extracellular space.
  • a major component of senile plaques is beta-amyloid (A-beta), a naturally secreted but insoluble peptide formed by cleavage of amyloid precursor protein (APP).
  • A-beta is a fragment close to the carboxyterminal domain of APP.
  • Neurofibrillary tangles are intraneuronal accumulations of filamentous material in the form of loops, coils or tangled masses. They are most abundantly present in parts of the brain associated with memory functions, such as the hippocampus and adjacent parts of the temporal lobe. See Robbins Pathologic Basis of Disease, Cotran et al., ⁇ .sup.th ed. (1999). Neurofibrillary tangles are commonly found in cortical neurons, especially in the entorhinal cortex, as well as in other locations such as pyramidal cells of the hippocampus, the amygdala, the basal forebrain, and the raphe nuclei.
  • Neurofibrillary tangles can also be found during normal aging of the brain, however, they are found in a significantly higher density in the brain of Alzheimer's disease patients, and in the brains of patients with other neurodegenerative diseases, such as progressive supranuclear palsy, postencephaltic Parkinson disease, Pick's disease, amylotrophic lateral sclerosis, etc. Robbins Pathologic Basis of Disease, Cotran et al., 6th ed. (1999), p.1330. Previous studies suggest that, among other things, neurofibrillary tangles may significantly contribute to the cognitive decline associated with the disease and also directly to neuronal cell death.
  • tau paired helical filaments
  • tau protein also referred to as "native tau”
  • Tau has an apparent molecular weight of about 55 kDa.
  • tau fragment The protease cathepsin D cleaves tau protein at neutral (cytoplasmic) pH resulting in tau fragments— one of which has a molecular weight of approximately 29 kDa (referred to by some authors as "tau fragment"). See, e.g., Bednarski & Lynch, J. Neurochem. 67:1846-1855 (1996); Bednarski & Lynch, NeuroReport 9:2089-2094 (1998). Both the tau protein and 29 kDa tau fragment can be phosphorylated. In a normal brain, the tau protein and tau fragment typically exist in an unphosphorylated, or dephosphorylated state.
  • both tau protein and tau fragment can be found in an abnormally phosphorylated state, a hyperphosphorylated state.
  • the 29 kDa tau fragment is a major component of neurofibrillary tangles.
  • Hyperphosphorylation impairs tau protein's ability to interact with microtubules.
  • Bednarski E, and Lynch G J Neurochem 67: 1846-55 (1996) cultured hippocampal slices with an inhibitor [N-CBZ-L-phenylalanyl-L-alanine-diazomethyl ketone (ZPAD)] of cathepsins B and L.
  • ZPAD N-CBZ-L-phenylalanyl-L-alanine-diazomethyl ketone
  • the ApoE4 isoform is a known risk factor for late- onset Alzheimer 's disease. Inhibition of cholesterol synthesis enhanced the phosphorylation of tau in dissociated cell cultures [ref. in (Sawamura, N., et al., J. Biol. Chem. 57: 1439-1443 (2001))]. Likewise, hyperphosphorylation of tau has been demonstrated in cell cultures prepared from NPC mutant mice (Sawamura, N., et al., J. Biol. Chem. 57:1439-1443 (2001)).
  • the present invention involves formulations of zinc (and more preferentially, gastroretentive sustained release zinc) and folic acid to reduce and stabilize the systemic and CSF levels of low molecular weight copper cysteine complexes (such as copper-homocysteine) that the present inventors recognize as a contributing factor to neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease and ALS, for example.
  • amyloid precursor protein APP
  • beta amyloid A ⁇
  • tau including the paired helical fragments (PHFs) and neurofibrillary tangle (NFTs)
  • beta secretase BACEl
  • apoE apolipoprotein E
  • apoE2, apoE3 and apoE4 differ in their ability to bind copper.
  • Epidemiological genetic studies indicate that presence of the apoE4 varient, having no cysteine residues at positions 112 and 158, increases the risk of AD, while apoE2, having two cysteine residues at positions 112 and 158, is considered to have protective benefit for Alzheimer's disease (as well as athlerosclerosis) as compared to the most common hum varient apoE3, which has only cysteine residue these positions.
  • homocysteine has been implicated with an increased risk of Alzheimer's disease, although until the disclosure contained herein, the effects homocysteine as a low molecular weight copper binding protein capable of delivering, maintaining and slowing clearance of toxic, loosely bound, and therefore exchangable "free" copper in the CNS has not been previously described.
  • elevated levels of cholesterol have been implicated with an increased risk of Alzheimer's disease.
  • elevated levels of oxidized cholesterol, 27S-hydroxy-cholesterol and/or 24S-hydroxy-cholesterol have been found both in the CNS and circulation and circulation of Alzheimer's patients (as well athlerosclerosis).
  • Neural Tube defects are major birth defects of the fetal brain or spine, and occurs when the neural tube (that later turns into the brain and spine) doesn't properly form, resulting in brain or spine damage. This occurs within the first few weeks a woman is pregnant, often before a woman knows that she is pregnant. Adequate intake of the B vitamin, folic acid by mothers prior to pregnancy has been shown to reduce the incidence of NTDs by up to 70% although the mechanism by which folic acid exerts this benefit has not yet been previously described. CDC, Folic Acid Now, CDC-NCEH99-0463, Nov. 2005. Spina bifida and anencephaly are two common types of NTDs.
  • the present invention provides formulations useful for lowering and maintaining steady systemic and CSF levels of free copper and iron which may be formulated and combined with one or more antioxidants, including, for example, vitamin C, vitamin E, QlO, omega 3 fatty acid, zinc-cysteine or combinations thereof.
  • one or more antioxidants including, for example, vitamin C, vitamin E, QlO, omega 3 fatty acid, zinc-cysteine or combinations thereof.
  • Acetylcholine Esterase Inhibitors are highly regarded clinical agents for treating and improving senile dementia such as Alzheimer type senile dementia, or cerebrovascular dementia, attention deficit hyperactivity disorder and schizophrenia.
  • donepezil hydrochloride (l-benzyl-4-[(5,6-dimethoxy-l-indanone)-2- yl]methylpiperidine hydrochloride) has been found to be useful as a acetylcholinesterase inhibitor in providing a desired pharmacological activity with minimum adverse side effects.
  • acetylcholinesterase inhibitors include rivastigmine (3-[l- (dimethylamino)ethyl]phenyl N-ethyl-N-methylcarbamate), metrifonate (dimethyl 2,2,2-trichloro-l-hydroxyethyl)phosphate), tacrine hydrochloride (1,2,3,4- tetrahydro-9-acridinamine), galanthamine hydrobromide, neostigmine, physostigmine etc.
  • An object of the present invention as further described herein includes formulations that combine acetyl-cholinesterase inhibitors with agents selected from the group of zinc, zinc-cysteine tetrathiomolybdate, gastroretentive sustained release zinc formulations and sustained release formulations of other essential trace metals such as, copper and iron.
  • NMDA receptor antagonists Excess activation of ionotropic glutamate receptors sensitive to N-methyl-D- aspartate (NMDA receptors) produces neuronal death and has been known to mediate various neurological diseases [Choi, Neuron 1:623-634 (1988)]. Glutamate, the excitatory neurotransmitter, is massively accumulated in brain subjected to hypoxic-ischemic injuries, which activates ionotropic glutamate receptors permeable to Ca 2+ and Na + and then causes neuronal death [Choi and Rothman, Annu Rev Neurosci 13:171-182 (1990)].
  • NMDA receptor antagonists possess therapeutic potentials to protect brain against hypoglycemia, hypoxia, and hypoxic-ischemic injuries.
  • NMDA receptor antagonists attenuate neuronal death following traumatic brain or spinal cord injuries [Faden, Lemke, Simon, and Noble. J.Neurotrauma. 5:33-45(1988); Okiyama, Smith, White, Richter, and Mclntosh. J.Neurotrauma. 14:211-222 (1997)].
  • Glutamate plays a central role in the induction and the propagation of seizures [Dingledine, McBain, and McNamara, Trends. Pharmacol. ScL 11:334-338 (1990); Holmes. Cleve.Clin.J.Med. 62:240-247 (1995)].
  • NMDA receptor antagonists were shown to act as anticonvulsants and antiepileptogenic drugs in various models of epilepsy [Anderson, Swartzwelder, and Wilson, J.Neurophysiol. 57:1-21 (1987); Wong, Coulter, Choi, and Prince. Neurosci.Lett.
  • ALS Amyotrophic lateral sclerosis
  • ALS is accompanied by degeneration of both upper and lower motor neurons and marked by neurogenic atrophy, weakness, and fasciculation. While the pathogenesis of ALS remains to be resolved, excitotoxicity has been expected to participate in the process of ALS.
  • ALS patients show increased levels of extracellular glutamate and defects in glutamate transport.
  • Administration of excito toxins mimicked pathological changes in the spinal cord of ALS patients [Rothstein. Clin.Neurosci.
  • NMDA receptors Antagonizing NMDA receptors appears to be applied to treat Parkinson's disease (PD).
  • MPTP l-methyl-4-phenyl-l,2,3,6-tetrahydropyridine
  • NMDA receptors protect dopaminergic neurons from the neurotoxin MPTP (l-methyl-4-phenyl-l,2,3,6-tetrahydropyridine) [Lange, Loschmann, Sofic, Burg, Horowski, Kalveram, Wachtel, and Riederer. Naunyn Schmiedebergs Arch. Pharmacol. 348:586-592 (1993); Brouillet and Beal. Neuroreport. 4:387-390 (1993)].
  • NMDA receptor antagonists also ameliorate levodopa-induced dyskinesia and thus can improve the therapeutic effects of levodopa [Papa and Chase, Ann.Neurol. 39:574-578 (1996); Marin, Papa, Engber, Bonastre, Tolosa, and Chase, Brain Res. 736:202-205 (1996)].
  • Two NMDA receptor antagonists, memantine and dextromethophan, have been proved beneficial in treating PD patients [Verhagen, Del Dotto, Natte, van den Munckhof, and Chase, Neurology 51:203-206 (1998); Merello, Nouzeilles, Cammarota, and Leiguarda. Clin.Neuropharmacol. 22:273-276 (1999)].
  • HD Huntington's disease
  • Another object of the present invention as further described herein includes formulations that combine NMDA antagonists, such as, memantine and flupirtine with agents selected from the group of zinc, zinc-cysteine, tetrathiomolybdate, gastroretentive sustained release zinc formulations and sustained release formulations of other essential trace metals such as, copper and iron.
  • NMDA antagonists such as, memantine and flupirtine
  • Free Radicals and Brain Diseases Free radicals are produced in degenerating brain areas following hypoxic- ischemia or traumatic brain and spinal cord injuries [Hall and Braughler, Free Radic.Biol.Med. 3:13-313 (1989); Anderson and Hall, Ann.Emerg.Med.
  • AD Alzheimers disease
  • Blockade of Zn 2+ translocation with Ca-EDTA attenuates neuronal loss following a transient forebrain ischemia or traumatic brain injury [Koh, Suh, Gwag, He, Hsu and Choi, Science 272: 1013-1016 (1996); Suh, Chen, Motamedi, Bell, Listiak, Pons, Danscher, and Frederickson, Brain Res. 852 :268-273 (2000)].
  • Yet another aspect of the present invention includes the use of gastroretentive sustained release formulations of zinc to increase intestinal, systemic and cerebral spinal fluid (CSF) levels of zinc and induce intestinal metallothienein and thereby reduce and/or maintain stable levels of loosely bound, "free" copper in the systemic circulation and CSF.
  • Cholesterol Sterols are structural lipids present in the membranes of all eukaryotic cells. These lipids are rigid and characterized by a four ring hydrocarbon steroid nucleus. Sterols are required not only to impart membrane fluidity, but also serve as the precursors for a variety of products with specific biological activities.
  • cholesterol an amphipathic sterol with a polar hydroxyl head group and nonpolar hydrocarbon body (the steroid nucleus)
  • Cholesterol is an essential molecule, playing a critical role in the structural integrity of cell membranes, a precursor for steroid hormones and serves as a precursor for bile acids. Cholesterol is synthesized in all organs but especially the liver from acetate and further obtained via dietary intake. Although cholesterol is a requisite molecule, high levels of blood cholesterol carried in the apoB containing lipoprotein or hypercholesterolemia has been implicated in atherosclerosis, heart attack, and stroke (Schultheis, 1990; Mitchell, 1990).
  • Hypercholesterolemia if not controlled, is one of several conditions that can lead to coronary artery disease. Coronary artery disease is the leading cause of death in the United States, accounting for approximately 600,000 deaths per year. Thus, the need exists for methods of treatment that can reduce cholesterol levels and methods to screen patients at risk for high cholesterol. Possible targets for treatment are transcription factors involved in cholesterol metabolism.
  • transcription factors involved in cholesterol metabolism are transcription factors involved in cholesterol metabolism.
  • nuclear receptors are ligand- activated transcription factors that govern aspects of every major developmental and metabolic pathway (reviewed in Kastner et al., 1995; Mangelsdorf et al., 1995). For example, the LXRs were first identified as "orphan" members of the nuclear receptor superfamily whose ligands and functions are unknown (Willy and Mangelsdorf, 1998).
  • the LXRs have recently been shown to be activated by a specific class of naturally occurring, oxidized derivatives of cholesterol, including 22(R)- hydroxycholesterol, 24(S)-hydroxycholesterol, and 24,25(S)-epoxycholesterol (Janowski et al., 1996; Lehmann et al., 1997). Oxysterols are concentrated in tissues where cholesterol metabolism and LXR expression are high, such as liver, brain, and placenta (Lavy et al., 1977; Spencer et al., 1985; Lutjohann et al., 1996).
  • LXRs function as heterodimers with the retinoid X receptors (RXRs), and thus, the RXR/LXR complex can be activated by both RXR ligands (i.e., rexinoids) and oxysterols (Teboul et al., 1995; Willy et al., 1995; Janowski et al., 1996).
  • RXR ligands i.e., rexinoids
  • oxysterols Teboul et al., 1995; Willy et al., 1995; Janowski et al., 1996.
  • Two LXR proteins (alpha and beta.) are known to exist in mammals. The expression of LXRalpha is restricted, with highest levels in the liver (hence, the name liver X receptor) and lower but significant levels in kidney, intestine, spleen, and adrenals (Apfel et al., 1994; Willy et al., 1995).
  • LXR.beta. expression is more widespread and has been found in nearly every tissue examined (Shinar et al., 1994; Song et al., 1994).
  • the pattern of expression of LXRs and their oxysterol ligands first suggested that these receptors may have a role in cholesterol metabolism.
  • Cholesterol has three essential metabolic fates in mammals: esterification (for transport or storage), and conversion into steroid hormones, or bile acids. Since steroid hormone synthesis is known to be governed by the orphan nuclear receptor steroidogenic factor- 1 (SF-I) (Parker and Schimmer, 1997), it is possible that LXRs are involved in bile acid synthesis (Janowski et al., 1996).
  • Cyp7a alpha. -hydroxylase
  • the Cyp7a promoter contains a functional LXR response element that can be activated by RXR/LXR heterodimers in an oxysterol- and retinoid-dependent manner (Lehmann et (al., 1997).
  • the formation of bile acids is one of two major pathways for the catabolism and excretion of cholesterol in mammals (Russell and Setchell, 1992).
  • LXRs may function as transcriptional control points in bile acid metabolism.
  • the RXR protein of RXR homo- and heterodimers has been observed to be regulated by 9-cis retinoic acid, which binds to the carboxy-teminus of RXR (Mangelsdorf and Evans, 1995).
  • RXR can form heterodimers with numerous other proteins in the nuclear receptor superfamily, including LXR. Depending on the receptor protein that dimerizes with RXR, and the ligands present, the resulting effects of the heterodimer on transcription can vary.
  • Synthetic retinoids have been found to selectively bind and activate RXRs (U.S. Pat. No. 5,780,676 and U.S. Pat. No. 5,455,265).
  • U.S. Pat. No. 6,835,866 describes the use of RXR-specific ligands, such as LG100268 to improve hepatic clearance of cholesterol.
  • RXR-specific ligands such as LG100268 to improve hepatic clearance of cholesterol.
  • the potential to modulate lipid concentrations in vivo, by targeting proteins of the nuclear hormone receptor superfamily with specific ligands may be useful in the treatment of various diseases related to lipid metabolism. For example, high blood cholesterol levels are associated with coronary disease. Lowering dietary cholesterol intake can significantly reduce cholesterol levels in most people.
  • cholestyramine and colestipol are resins that bind bile acids in the intestinal tract, causing the liver to increase its production of bile acids and thus lower the cholesterol levels, by converting cholesterol into bile acid. Nicotinic acid, gemfibrozil, probucol, as well as statins such as atorvastatin, lovastatin, pravastatin are commonly used to lower blood lipid levels.
  • bile acids such as, ursodeoxycholic acid (UDCA) 17.beta.-(l- methyl-S-carboxypropyljetiocholane-S. alpha., 7.beta.diol. are known to improve biliary function. No major toxicity is known to be associated with UDCA (W. H. Bachrach et al., Dig. Dis. ScL, 30, 642 (1985)).
  • salts and esters of UDCA include nontoxic esters of the free hydroxyl groups of UDCA with (Ci -C 4 )alkanoic acids such as formic, acetic or propionic acid, phosphate esters of the OH groups, (Ci -C 4 )alkyl esters of the free (C 24 ) carboxylic acid group and nontoxic alkali metal, ammonium or amine salts of the free carboxylic acid moiety.
  • This ester and salts can be readily prepared from free UCDA by methods well known to the art. At least the diformate and diacetate esters are known compounds. See, The Merck Index (1 lth ed. 1989) at 1556.
  • UDCA is commercially available in 300 mg hard gelatin capsules as ACTIGALL® from Summit Pharmaceuticals, Summit, NJ. , and is prescribed for gallbladder stone dissolution.
  • the present invention in one aspect is based in part upon the unique finding that circulating levels of loosely bound or so-called "free" copper and iron in the systemic circulation and CSF are subject to substantial intraday peak fluctuations which are exacerbated by bolus administration of copper and iron in soluble form such as that contained in ordinary tap water as well as upon dissolution of immediate release copper or iron supplements.
  • a feature of the present invention as further described herein includes formulations that combine agents capable of improving hepatic and biliary clearance functions, such as, statins, including atorvastatin, RXR specific ligands, such as LG1002688, with agents selected from the group of zinc, zinc-cysteine, tetrathiomolybdate, gastroretentive sustained release zinc formulations and sustained release formulations of other essential trace metals such as, copper and iron so as to improve hepatic incorporation and clearance of free copper and iron in the systemic circulation and CSF of persons having potentially impaired hepatic and biliary function to treat or prevent neurodegenerative disorders, such as, Alzheimer's disease, mild cognitive impairment, Parkinson's disease, ALS and atherosclerosis caused by elevated levels and fluctuating levels of free copper and iron in the systemic circulation and CSF.
  • statins including atorvastatin, RXR specific ligands, such as LG1002688
  • such formulations may reduce the potential for hypocupremia and anemia through chroic zinc administration by the addition to such formulations of sustained release formulations containing copper, iron and/or other essential trace metals which preferentially will be bound to certain copper and iron binding excipients, such as whey, plant fibers, metallotheionein, dried milk or infant formula.
  • Parkinson' s Disease Parkinson's disease is a common neurodegenerative disorder and was first described by James Parkinson in 1817. The four primary diagnostic signs of the illness are resting tremor, bradykinesia, muscular rigidity and postural instability.
  • PD dopaminergic neurons
  • Gibb, W., et al., J. Neurol. Neurosurg. and Psych., 51:745-52 (1988) PD is characterized by the formation of Lewy Bodies and death of dopaminergic neurons. [Adams D. et al., Principles of Neurology, 874-880, 3rd Edition, McGraw-Hill, N.Y., (1985)].
  • the neuropathological hallmark of PD is the LewyBody.
  • Lewy Bodies are intracytoplasmic inclusions that occur in degenerating neurons which are composed of a dense core of filamentous and granular material surrounded by radical oriented filaments that have a diameter of 10-20 nm [Goedert, 20 M., et al., Curr. Op. Neurobio. 8:619-32 (1999)].
  • the causes of PD are not known and there has been vigorous debate over the relative roles of genetics and environmental factors [Tanner, C, et al., JAMA, 281:341-6 (1999)].
  • Alpha-Synuclein is a ubiquitous protein that shares significant physical and functional homology to the protein chaperone, 14-3-3, and is particularly abundant in the brain (Ostrerova N. et al., J. Neurosci., 19:5782 (1990); Clayton D. et al., TINS 21:249 (1998)].
  • Alpha-Synuclein is normally phosphorylated at serines 87 and 129. (Okochi M. et al., J. Biol. Chem., 275:390 (2000)]. Recent studies showed that mutations in alpha-synuclein can cause familial PD and that alpha-synuclein accumulates in LewyBodies.
  • Lewy Bodies could either be inert tombstone markers that occur in response to free radical damage, or they might be toxic agents that harm the cell. Examples of both situations exist in the literature. Aggregated amyloid-beta (beta.) is toxic to neurons, while lipofuscin appears to be innocuous to cells (Behl, C, et al., Cell 77:817-27 (1994)]. The Huntington's protein presents an intermediate situation where the toxicity associated with Huntington's appears to precede aggregation, and aggregation of Huntington's protein might even be protective [Saudou, F., et al., Cell 95:55-66 (1998)].
  • mice over-expressing alpha-synuclein show an age-related loss of dopaminergic terminals and motor impairment, which could be indicative of toxicity [Masliah, E. et al., Science, 287:1265-1269 (2000)].
  • Oxidative stress is thought to contribute to PD because dopamine, which is a strong free radical generator, is the principle neurotransmitter in the substantia nigra [Chiueh, C, et at., Adv. Neurol., 60:251-8 (1993); Jenner, P. et al., Ann. Neurol., 25 44:S72-84 (1998)].
  • iron which also stimulates free radical production, accumulates in the substantia nigra with age [Jenner, P., et al., Ann. Neurol., 44:S72-84 (1998)].
  • Nonalcoholic steatohepatitis involves the development of histologic changes in the liver that are comparable to those induced by excessive alcohol intake but in the absence of alcohol abuse. Macrovesicular and/or microvesicular steatosis, lobular and portal inflammation, and occasionally Mallory bodies with fibrosis and cirrhosis characterize NASH. NASH is also commonly associated with hyperlipidemia, obesity, and type II diabetes mellitus.
  • nonalcoholic steatohepatitis can be used to describe those patients who exhibit these biopsy findings, coupled with the absence of (a) significant alcohol consumption, (b) previous surgery for weight loss, (c) history of drug use associated with steatohepatitis, (d) evidence of genetic liver disease or (e) chronic hepatitis C infection. See, J. R. Ludwig et al., Mayo Clin.
  • Steatohepatitis is now considered an important cause of end-stage liver disease and may be the cause of an unknown number of cases of clyptogenic cirrhosis. See E. E. Powell et al, cited above. Unfortunately, once cirrhosis is established, the only therapeutic modality available is orthotopic liver transplantation. Thus, effective therapy for nonalcoholic steatohepatitis is clearly needed.
  • Multiple Sclerosis Elemental copper and is also known to play a role in the formation of myelin. Antigenic forms of myelin are also associated with the T-cell mediated lesions and myelin destructiion that is characteristic of multiple sclerosis. Autism Metabolic copper dysfunction has also been implicated in autism. Chuahan, A et. al, Life Sci.
  • yet another object of the present invention includes the treatment of autism with gastroretentive and/or sustained release formulations of zinc, copper and iron and other trace metals so as to maintain a targeted and steady level of free copper and iron in the systemic circulation and CSF of autistic children.
  • the present invention in one aspect provides improved pharmaceutical compositions, kits and methods to improve the means to induce, monitor and safely maintain a state of copper malabsorption for extended periods in a person or animal in need of reducing, managing or maintaining low levels of free, unbound or loosely-bound copper, including, but not limited to, the indications of Wilson's disease, Alzheimer's disease, atherosclerosis, autoimmune diseases, oxidative stress, geriatric-related impaired copper excretion, liver disease, neurodegenerative disorders such as ALS, Parkinson's disease, and multiple sclerosis, as well as diseases associated with elevated levels of cuproproteins, such as schizophrenia. Also disclosed are specially formulated pharmacants, kits and dosing regimens intended to reduce the need to monitor patients for signs of hypercupronemia.
  • the present invention in one aspect provides pharmaceutical and over- the-counter products designed and formulated specifically to block absorption of copper and copper ions from the gastrointestinal tract, while also limiting the systemic bioavailability of such blocking agents.
  • Novel Zinc Containing Formulations to Induce and Maintain a Targeted and Steady State Copper Balance Zinc acetate formulated as an immediate release 25 mg and 50 mg capsule was developed and clinically tested as a maintenance therapy for the treatment of Wilson's disease.
  • the recommended dosage for Wilson's disease patients is 50 mg taken three times a day (t.i.d.) at least one hour before meals and beverages (other than water) and at least one hour after meals and beverages (other than water).
  • Zinc blocks the intestinal absorption of copper from the diet and the re- absorption of endogenously secreted copper such as that from saliva, gastric juice and bile. Zinc induces the production of metallothionein in the enterocyte, a protein that binds copper and thereby prevents its serosal transfer into the blood. The bound copper is then lost in the stool following desquamation of the intestinal cells.
  • Gastroretentive Agents that are Useful for the Maintenance of Copper Malabsorption
  • Various pharmaceutical controlled release drug delivery systems with prolonged gastric retention time have been described in the literature. These involve different technologies. The advantages of using drug delivery systems are many. Of major importance in controlled drug therapy is improved efficiency in treatment. Controlled drug therapy reduces the required frequency of administration, and single doses at periodic intervals are sufficient, resulting in improved patient compliance.
  • Current formulations of zinc acetate require dosing of at least two to three times per day in order to maintain a state of copper malabsorption.
  • the present invention provides a benefit in diseases other than Wilson's disease, such as Alzheimer's disease, because it helps to achieve a comparable or greater level of copper malabsorption with an equal or lower amount of systemic zinc exposure.
  • copper has been found to be bound to amyloid beta plaques in the brains of autopsy-confirmed Alzheimer's disease patients.
  • the present invention by providing a means to block copper absorption in the intestines while minimizing serum levels of zinc, reduces the uncertainty regarding the relationship between Alzheimer's disease and zinc.
  • Zinc may play a beneficial role in the serum and CNS given its status as an anti-oxidant and zinc found in Alzheimer's plaques may not be causally related as in the case of copper.
  • a gastroretentive pharmaceutical dosage form can be developed utilizing a number of alternative gastroretentive systems. After oral administration, such gastroretentive dosage form should retain in the stomach and release zinc acetate in a controlled and prolonged manner.
  • gastroretentive dosage forms are floating dosage forms and dosage forms that expand, swell or unfold in the stomach.
  • the rationale for developing expandable drug delivery systems is based on the nature of the pyloric antrum that, by means of antiperistaltic motion, retropels large bodies away from the pylorus back to the fundus and body of the stomach, thus prolonging their gastric retention time (GRT).
  • Such dosage forms should preferably be designed to undergo biodegradation or disintegration to enable their evacuation from the stomach.
  • U.S. Pat. No. 4,207,890 discloses a drug dispensing device which comprises a collapsed, expandable imperforate envelope, made of a non-hydratable, body fluid and drug-permeable polymeric film, which contains the drug, and an expanding agent also contained within the polymeric envelope which, when in contact with body fluids, causes the envelope to expand to a volume such that the device is retained in the stomach.
  • 4,434,153 describes a device comprised of a matrix formed of a hydrogel that absorbs and imbibes fluid from the stomach, expands, and swells in order to retain in the stomach for an extended period of time, and a plurality of tiny pills dispersed throughout the matrix, having a drug-containing core and a fatty acid and wax wall surrounding the core.
  • a significant disadvantage of the devices of the prior art publications cited above is that they appear to ignore natural contractions of the stomach which may contribute to a rapid diminishing of size, leading to early removal of the device from the stomach. These devices lack the strength required to withstand the natural mechanical activity that includes contractions of the stomach.
  • 4,767,627, 4,735,804 and 4,758,436 present dosage forms of various geometries: continuous solid stick; tetrahedron; planar disc; multi-lobed flat device; and ring.
  • the devices are compressible to a size suitable for swallowing, and are self-expandable to a size which prevents passage through the pylorus. They are sufficiently resistant to forces of the stomach to prevent rapid passage through the pylorus for a pre-determined period of time and erode in the presence of gastric juices.
  • the devices are homogenous, namely they contain the same polymer constitution in different areas of the device.
  • 4,735,804 is homogenous in its four lobes, which are attached to each other by a polymeric matrix.
  • Zinc acetate is incorporated into the device as a liquid solution or suspension, which may necessitate the addition of mentioned preservatives or buffering agents.
  • the controlled release zinc acetate module may be tethered or glued to the device.
  • U.S. Pat. Nos. 5,002,772 and 5,443,843 disclose an oral drug delivery system having a delayed gastrointestinal transit, which releases the drugs contained therein in a controlled manner and which in their expanded form resist gastrointestinal transit.
  • These delivery systems comprise one or more retention arms as a non- continuous compressible element, and an attached controlled release drug-containing device.
  • the preferred configuration is a coil or a spiral.
  • U.S. Pat. Nos. 5,047,464 and 5,217,712 describe a system comprising bio- erodible, thermoset, covalently cross-linked, poly(ortho) ester polymers, which expand from a compressed state upon delivery thereof.
  • the acidic environment of the stomach eventually results in the degradation of the polymers within the system, thus permitting its removal from the stomach.
  • the system is characterized by high resiliency.
  • 6,685,962 describes a gastroretentive drug delivery system for the controlled release of an active agent in the gastrointestinal tract which comprises: (a) a single- or multi-layered matrix comprising a polymer that does not retain in the stomach more than a conventional dosage form selected from (1) degradable polymers that may be hydrophilic polymers not instantly soluble in gastric fluids, enteric polymers substantially insoluble at pH less than 5.5 and/or hydrophobic polymers and mixtures thereof; (2) non-degradable polymers; and any mixtures of (1) and (2); (b) a continuous or non-continuous membrane comprising at least one polymer having a substantial mechanical strength; and (c) a drug; wherein the matrix when affixed or attached to the membrane prevents evacuation from the stomach of the delivery system for a period of time.
  • a conventional dosage form selected from (1) degradable polymers that may be hydrophilic polymers not instantly soluble in gastric fluids, enteric polymers substantially insoluble at pH less than 5.5 and/or hydrophobic polymers and mixture
  • a pharmaceutical composition can be formulated for oral, intravenous, intramuscular, subcutaneous, or inhalation administration as well as by other routes (i.e. enema, intranasal, intrathecal, etc).
  • Advantages of orally administered pharmaceuticals include rapid therapeutic effect and patient convenience. It is known in the art to orally administer a pharmaceutical in order to provide a direct effect on a target site within the gastrointestinal tract, as opposed to providing a therapeutic effect by absorption of the active ingredient of the pharmaceutical composition into the patient's circulatory system (i.e. antacids, laxatives).
  • the controlled gastric retention of solid dosage forms of a pharmaceutical can be achieved by the mechanisms of mucoadhesion, flotation, sedimentation, expansion, or by the simultaneous administration of pharmacological agents which delay gastric emptying.
  • Mucoadhesion is the process whereby synthetic and natural macromolecules adhere to mucosal surfaces in the body. If these materials are then incorporated into pharmaceutical formulations, drug absorption by mucosal cells can be enhanced or the drug released at the site for an extended period of time.
  • synthetic polymers such as the chitosans, carbopols and carbomers
  • the mechanism of bio/mucoadhesion is the result of a number of different physicochemical interactions.
  • Biological bio/mucoadhesives such as plant lectins, show specific interactions with cell surfaces and mucin and are seen as the "second generation" bioadhesives.
  • mucoadhesion acts to impart to orally administered dosage forms the ability to resist the strong propulsion forces of the stomach wall.
  • the continuous production of mucous by the gastric mucosa to replace the mucous which is lost through the peristaltic contractions and the dilution of the stomach content can be overcome by use of mucoadhesion as a gastroretentive force.
  • Mucoadhesive nanoparticulate systems including liposomes and polymeric nanoparticles
  • Mucoadhesive ability can be conferred on particulate systems by coating their surface with mucoadhesive polymers such as chitosan and Carbopol. The feasibility of such surface modification has been confirmed by measuring the zeta potential. Evaluation procedures include a particle counting method using a Coulter counter for polymer-coated liposomes.
  • Mucoadhesive nanoparticles have been used for the oral administration of peptide drugs, and have been shown to be more effective with a more prolonged action as compared to non-coated systems (Takeuchi H., et al.).
  • Mucoadhesive nanoparticulate systems for peptide drug delivery offer several advantages over traditional dosage forms including the ability to optimize the therapeutic effects of a drug by controlling its release into the body. It has been shown that various types of poly(acrylic acid) (PAA) hydrogels are able to inhibit the hydrolytic activity of gastrointestinal enzymes, such as trypsin, resulting in an increase of the bioavailability of the drug. Acrylic-based polymers can be used for the attachment of mucoadhesive delivery systems to the mucosa.
  • PAA poly(acrylic acid)
  • Polymer hydrogels modified by grafting mucophilic copolymers such as poly(ethylene glycol) (PEG) onto the back- bone chains of the polymer can promote the adhesive process. This is due to the ability of these grafted chains to diffuse from the network to the mucous layer.
  • Films of P(AA-g-EG) can be synthesized by using UV-initiated free-radical solution polymerization.
  • Different types of hydrogels can be synthesized with varying molar feed ratio of AA to PEG.
  • the polymer hydrogels are characterized by mucoadhesion in order to quantify the effects of the PEG grafted chains on mucoadhesion.
  • the bioadhesive bond strength can be determined using a tensile apparatus, and the work of adhesion thereby calculated.
  • Hydrogels containing 40% AA and 60% PEG (40:60 AA/EG) can exhibit the highest mucoadhesion. These results can be attributed to the synergistic effects of both monomers.
  • AA functional groups can permit the polymer to form multiple hydrogen bonds as well as to swell to a large degree.
  • PEG tethers acted as mucoadhesive promoters. They penetrated into the mucosa and bridged the base hydrogel and the mucus. These results can also be interpreted in terms of the Huang-Peppas models (2002) of surface coverage and chain length effects in mucoadhesion.
  • flotation as a retention mechanism requires the presence of liquid on which the dosage form can float, and it also presumes that the patient remains in an upright posture during the GRI, because in a supine position the pylorus is located above the stomach body and allows the accelerated emptying of floating material.
  • flotation can be a basis principle for gastric retention of an oral formulation. Sedimentation has been successfully used as a retention mechanism for pellets which are small enough to be retained in the rugae or folds of the stomach body near the pyloric region, which is the part of the organ with the lowest position in an upright posture. Dense pellets (of greater than approx.
  • Elemental zinc has a density of 7.165 g/cm 3 which is 4.365 g/cm 3 greater than the 2.8 g/cm 3 threshold density necessary for gastroretention.
  • Zinc's high specific density of 7.165 g/cm 3 creates the opportunity to utilize up to approximately 60.9% of each pellet to incorporate other desirable, less dense materials to improve the function, safety, tolerability and effectiveness of the gastroretentive zinc pellets. Accordingly, the present invention describes an enteric-coated zinc pellet of sufficiently small size to become trapped in the rugae.
  • Such an enteric coating provides the advantage of avoiding the potential for irritation to the stomach wall as a result of direct contact with the trapped zinc pellet.
  • the enteric coating may be pH dependent and selected to prevent degradation of the coating and release of the zinc while the pellet is still in the low pH conditions of the stomach and/or pyloric region (pH of 1.2-3.5).
  • the coating instead begins to degrade and release zinc in the substantially higher pH of the duodenum (pH of approx. 4.6-6.0), jejunum (pH of approx. 6.3-7.3) and/or colon or rectum (pH of approx.
  • the pellets may be contained within a swallowable capsule that rapidly dissolves and releases the pellets upon entry into the stomach.
  • the capsules may easily be varied according to the total amount of zinc contained, as well as by the mixture of number and type of pellets contained, to best suit the habits and dosing preferences of the patient and to provide greater confidence that the target "free” or "serum” copper will remain within the desired range. Expansion has been shown to be a potentially reliable retention mechanism.
  • Several devices have been described in the art which comprise features which extend, unfold or are inflated by carbon dioxide generated within the devices after administration.
  • the copper bound to proteins contained in foodstuffs will likely be processed by the intestines in a more deliberate, enzyme- specific manner than the free, unbound, solubilized, potentially ionic copper contained in drinking water.
  • a much greater percentage of the copper contained in drinking water will most likely be absorbed as a bolus because no digestion to free it from a food-legand complex is required.
  • the intestinal cells will pass their bolus on the blood, where it is primarily loosely bound by albumin.
  • bolus effect may allow a significant part of the copper to bypass the liver, i.e., bypassing the intestines' evolutionary copper absorption and regulatory apparatus, and be picked up by the brain.
  • Copper in drinking water could enter the serum, bind to small peptides, and cross the blood barrier into the central nervous system, bypassing the normal blood brain barrier regulation of copper transport and homeostasis. Free or loosely bound copper in the CNS and the serum are believed to be at equilibrium.
  • copper absorbed through drinking water is most likely absorbed in a bolus fashion as compared to copper absorbed through foods. The anticipated result is a much higher peak free copper concentration in the CNS as compared to the daily average.
  • Such a peak free copper concentration in the CNS would presumably upregulate the copper binding proteins/ protective mechanisms such as APP, amyloid beta and tau.
  • the copper molecules could also be in the ionic or cupric form and therefore of greater toxic potential. It is an object of the present invention to provide continuous protection for copper-sensitive individuals, such as Wilson's disease, Alzheimer's disease, dementia and elderly patients, from the toxic effects of peak serum and CNS free copper concentrations by administration of special formulations of one or more continuous release copper malabsorption agents, such as zinc and ascorbic acid, and thereby reduce the effects associated with toxic free copper, especially the upregulation of protective cuproproteins in the CNS.
  • the gastroretentive pill or capsule utilizes a combination of different gastroretentive mechanisms to assure the broadest and least variable protection.
  • capsules may be formulated with floating zinc salt-containing microparticles, mucoadhesive microparticles, mucoadhesive high density zinc-containing pellets, mucoadhesive high density enteric coated zinc-containing pellets, expanding gastroretentive systems containing zinc salts, as well as immediate release and/or non-gastroretentive zinc powder or enteric coated zinc microparticles.
  • Such formulations will aid the prescribing doctor in estimating a recommended daily dosage to achieve a certain level of copper protection or copper malabsorption.
  • Zinc Formulations of zinc acetate available as an FDA approved form, such as Galzin are associated with poor compliance on the part of patients due to gastric irritability, which is believed to be associated with the ionic nature of zinc.
  • Stomach irritability is associated with current formulations in an estimated 10% of patients. Since copper-containing liquids, such as water, or the copper contained in foods, do not begin to absorbed in the human body until they reach the small intestines, it is not necessary to induce expression of metallothienein in the stomach in order to maintain a state of copper malabsorption.
  • immediate release formulations of zinc that are intended to induce a state of copper malabsorption also unnecessarily increase the systemic circulating levels of zinc and to other organ systems, such as the brain.
  • Excess levels of zinc in the brain and central nervous system are implicated in certain neurodegenerative disorders in which excess copper is also implicated. Examples of such disorders include, but are not limited to, Alzheimer's disease, Parkinson's disease and amyotrophic later sclerosis (ALS).
  • an enteric coated formulation of zinc so that the zinc-containing pill, capsule, tablet (or microparticles contained therein) does not begin to release the content of zinc until after it has fully transited the esophagus and stomach.
  • a typical enteric coating may be a polymeric material.
  • Preferred enteric coating materials comprise bioerodible, gradually hydrolysable and/or gradually water-soluble polymers.
  • the "coating weight,” or relative amount of coating material per capsule generally dictates the time interval between ingestion and drug release. Any coating should be applied to a sufficient thickness such that the entire coating does not dissolve in the gastrointestinal fluids at pH below about 5, but does dissolve at pH about 5 and above.
  • any anionic polymer exhibiting a pH-dependent solubility profile can be used as an enteric coating in the practice of the present invention to achieve delivery of the active drug to the lower gastrointestinal tract.
  • the selection of the specific enteric coating material depends on the following properties: resistance to dissolution and disintegration in the stomach; impermeability to gastric fluids and drug/carrier/enzyme while in the stomach; ability to dissolve or disintegrate rapidly at the target intestine site; physical and chemical stability during storage; non-toxicity; ease of application as a coating (substrate friendly); and economical practicality.
  • Suitable enteric coating materials include, but are not limited to: cellulosic polymers such as hydroxypropyl cellulose, hydroxyethyl cellulose, hydroxypropyl methyl cellulose, methyl cellulose, ethyl cellulose, cellulose acetate, cellulose acetate phthalate, cellulose acetate trimellitate, hydroxypropylmethyl cellulose phthalate, hydroxypropyhnethyl cellulose succinate and carboxymethylcellulose sodium; acrylic acid polymers and copolymers, preferably formed from acrylic acid, methacrylic acid, methyl acrylate, ammonio methylacrylate, ethyl acrylate, methyl methacrylate and/or ethyl methacrylate (e.g., those copolymers sold under the tradename "Eudragit”); vinyl polymers and copolymers such as polyvinyl pyrrolidone, polyvinyl acetate, polyvinylacetate phthalate, vinylacetate crotonic acid copoly
  • enteric coating materials for use herein are those acrylic acid polymers and copolymers available under the tradename "Eudragit” from Rohm Pharma (Germany).
  • the Eudragit series E, L, S, RL, RS and NE copolymers are available as solubilized in organic solvent, as an aqueous dispersion, or as a dry powder.
  • the Eudragit series RL, NE, and RS copolymers are insoluble in the gastrointestinal tract but are permeable and are used primarily for extended release.
  • the Eudragit series E copolymers dissolve in the stomach.
  • the Eudragit series L, L-30D and S copolymers are insoluble in stomach and dissolve in the intestine, and are thus most preferred herein.
  • a particularly suitable methacrylic copolymer is Eudragit L, particularly L- 3OD and Eudragit 100-55.
  • Eudragit L-30D the ratio of free carboxyl groups to ester groups is approximately 1: 1.
  • the copolymer is known to be insoluble in gastrointestinal fluids having pH below 5.5, generally 1.5-5.5, i.e., the pH generally present in the fluid of the upper gastrointestinal tract, but readily soluble or partially soluble at pH above 5.5, i.e., the pH generally present in the fluid of lower gastrointestinal tract.
  • Eudragit S Another particularly suitable methacrylic acid polymer is Eudragit S, which differs from Eudragit L-30D in that the ratio of free carboxyl groups to ester groups is approximately 1:2.
  • Eudragit S is insoluble at pH below 5.5, but unlike Eudragit L-30D, is poorly soluble in gastrointestinal fluids having a pH in the range of 5.5 to 7.0, such as in the small intestine.
  • This copolymer is soluble at pH 7.0 and above, i.e., the pH generally found in the colon.
  • Eudragit S can be used alone as a coating to provide drug delivery in the large intestine.
  • Eudragit S being poorly soluble in intestinal fluids below pH 7, can be used in combination with Eudragit L-30D, soluble in intestinal fluids above pH 5.5, in order to provide a delayed release composition which can be formulated to deliver the active agent to various segments of the intestinal tract.
  • the enteric coating provides for controlled release of the active agent, such that drug release can be accomplished at some generally predictable location in the lower intestinal tract below the point at which drug release would occur without the enteric coating.
  • the enteric coating also prevents exposure of the hydrophilic therapeutic agent and carrier to the epithelial and mucosal tissue of the buccal cavity, pharynx, esophagus, and stomach, and to the enzymes associated with these tissues.
  • the enteric coating therefore helps to protect the active agent and a patient's internal tissue from any adverse event prior to drug release at the desired site of delivery.
  • the coated capsules of the present invention allow optimization of drug absorption, active agent protection, and safety.
  • the coating can, and usually does, contain a plasticizer to prevent the formation of pores and cracks that would permit the penetration of the gastric fluids.
  • Suitable plasticizers include, but are not limited to, triethyl citrate (Citroflex 2), triacetin (glyceryl triacetate), acetyl triethyl citrate (Citroflec A2), Carbowax 400 (polyethylene glycol 400), diethyl phthalate, tributyl citrate, acetylated monoglycerides, glycerol, fatty acid esters, propylene glycol, and dibutyl phthalate.
  • a coating comprised of an anionic carboxylic acrylic polymer will usually contain approximately 10% to 25% by weight of a plasticizer, particularly dibutyl phthalate, polyethylene glycol, triethyl citrate and triacetin.
  • the coating can also contain other coating excipients such as detackifiers, antifoaming agents, lubricants (e.g., magnesium stearate), and stabilizers (e.g., hydroxypropylcellulose, acids and bases) to solubilize or disperse the coating material, and to improve coating performance and the coated product.
  • the coating can be applied to the capsule using conventional coating methods and equipment. For example, an enteric coating can be applied to a capsule using a coating pan, an airless spray technique, fluidized bed coating equipment, or the like. Detailed information concerning materials, equipment and processes for preparing coated dosage forms may be found in Pharmaceutical Dosage Forms: Tablets, eds. Lieberman et al.
  • an object of the present invention is directed towards achieving and maintaining a healthy level of copper status in elderly or geriatric patients through the restoration of normalized hepatic copper excretion.
  • the present invention of normalizing hepatic excretion of copper has the important advantage of limiting the necessity to regularly monitor the copper status of each patient to assure that copper status remains within the targeted range, thereby avoiding copper toxicosis and hypercupronemia.
  • Agents that are useful for improving the excretion of copper in elderly patients include ursiodiol, anti-inflammatory agents and antifibrotics.
  • Depot Injection Formulations of Copper-Chelating, Copper-Complexing and Copper-Blocking Agents An alternative to oral delivery of agents that reduce body copper is by means of a subcutaneous injection or subcutaneous implant. Liposomes have proven to be versatile carriers for the delivery of drugs.
  • Multivesicular liposomes can be formulated with zinc to release in the body over the course of a month or more. As a result of their larger size (median diameter typically 10-30 ⁇ m), these multivesicular liposomes are not rapidly cleared by macrophages and can act as a drug depot, providing slow release of drugs for up to or more than one month. It is anticipated that such a depot formulation will improve compliance in Alzheimer' s patients and nevertheless readily induce metallotheionein in the intestines suffcient to block absorption of copper from the intestines.
  • subcutaneous implants have been developed and utilized to deliver a drug of interest for up to a year or more.
  • Such implants generally utilize polymer matrices that incorporate the drug of interest and permit its release in a time-dependent manner.
  • Osmotic pumps have also been utilized in a similar fashion.
  • An implant formulated with zinc could achieve easy compliance, especially in the case of Alzheimer's patients, and nevertheless readily induce metallothionein in the intestines sufficient to block absorption of copper from the intestines.
  • a transdermal patch could accomplish the same objectives as an implant and achieve steady state zinc in the blood stream.
  • a copper-blocking agent such as zinc will have the added benefit of reducing the peaks and throghs of loosely bound copper in the serum and other body compartments, thereby reducing the induction of copper transport proteins, such as, amyloid precursor protein, amyloid beta, and tau by neuronal cells at time points in which levels of intracellular copper are high. Accordingly, it is anticipated that, beyond the convenience of administration, such systems will have the added therapeutic benefit of reducing the total production of pathogenic copper transporters as compared to non-sustained release approaches, delivering a comparable average dose of zinc or other copper blocker, copper chelator or copper complexor.
  • an object of the present invention is to utilize formulations including ascorbic acid to block the absorption of copper from the gastrointestinal tract.
  • Diseases Associated with Hyperhomocysteinemia Elevated levels of serum homocysteine, a copper-binding protein having a molecular weight of 135 Daltons, have been associated with Alzheimer's disease, atherosclerosis and schizophrenia (the Framingham Study, NEJM 2003).
  • homocysteine represents an opportunistic pool of a circulating, low molecular weight, copper-binding protein that serves to maintain an elevated level of toxic, loosely bound copper that is also free to cross the blood brain barrier.
  • an object of the present invention to provide a method of treating schizophrenia and cardiovascular or other disease identified by elevated levels of homocysteine through the administration of a copper chelator, copper complexor or copper malabsorption agent, including zinc and gastroretentive formulations of zinc and zinc salts, and thereby reduce the circulating level of copper-bound homocysteine in the serum and CNS.
  • An object of the present invention is to utilize zinc-cysteine complexes, such as zinc-monocysteine described by Newsome in U.S. Pat. No. 6,586,611, as copper malabsorption agents for the treatment of Wilson's disease, schizophrenia, athlerosclerosis and neurodegenerative diseases associated with elevated levels of free copper, including Alzheimer's disease, ALS and Parkinson's disease.
  • intraday fluctuation of free copper in the CNS could help explain the contradictory conclusions reached by different groups as to whether a goal of Alzheimer's disease treatment should be the reduction of copper intake or copper supplementation. While free copper levels in the CNS and resulting intraneuronal free copper are elevated, it is likely that proteins capable of binding and precipitating intraneuronal free copper, such as, APP and the cleaved transmembrane peptide A ⁇ would be upregulated resulting in the precipitation of A ⁇ bound copper into the extraneuronal CSF.
  • proteins capable of binding and precipitating intraneuronal free copper such as, APP and the cleaved transmembrane peptide A ⁇ would be upregulated resulting in the precipitation of A ⁇ bound copper into the extraneuronal CSF.
  • the inventors attribute such free copper fluctuations to rapid influx copper containing water via the small intestines, the portal vein, serum plasma, crossing the blood brain barrier into the cerebral spinal fluid and brain.
  • solubilized copper such as that ordinarily found in tap water, exists in a free, unbound form and may rapidly enter the systemic circulation and CSF as a bolus shortly following influxes of water in the intestines due to the first pass effect.
  • Such copper influxes have been shown to be increased in the presence of sodium (Wapnir) and thereby bypass or overwhelm the copper regulatory mechanisms of intestinal epithelia and liver by virtue of their inability to respond in a timely fashion via the upregulation of intestinal metallotheionein and hepatic ceruloplasmin.
  • Water influxes of solubilized copper have been shown by Wapnir to be exacerbated in the presence of sodium thereby potentially implicating intestinal sodium channels as a potential entry point of solubilized copper in the portal vein and hepatic circulation. Copper bound to chylomicrons may also enter into the lymphatic circulation finally draining into the thoracic duct.
  • the beads are subsequently separately washed with distilled water and the eluted solution of each is measured and quantified for total radioactivity by means of standard protocols measuring total radioactivity indicating the total amount of Cu 4 present in such elution on an absolute and related on a percentage basis to both the total solution volume as well as quantity of ceruloplasmin, albumin or other proteins.
  • the results show both a higher average level of peak Cu 4 levels and AUC of Cu 4 in the albumin and other protein elutions in the Alzheimer's and Parkinson's patients compared to aged matched controls and young patients, as well as when age matched controls are compared to younger patients.
  • the results of the examples above could be reproduced by substituting a Cu 4 supplement as described by Bayer with Cu 64 drinking water previously described. Since the immediate release formulation described by Bayer would also result in elevated peak and AUC levels of free copper in the systemic circulation and CSF patients.
  • One cohort of 24 Alzheimer's patients are treated once a day for one to three months with an immediate release copper supplement as described by Bayer TA containing 2mg of copper.
  • a second cohort of 24 Alzheimer's patients are treated with once a day for the same period with a sustained release formulation containing the same quantity of copper either also as a salt or preferably bound to a natural copper binding carrier such as, metallotheionein, fiber, whey or casein. All patients abstain from copper containing drinking water during the study period and efforts are made to balance the groups based upon approximate daily intake of dietary copper-containing foods as well as use of cholesterol lowering agents and other medications.
  • Serial serum samples are taken at least 12 hours away from food every week at points alternately within 1-3 hours following daily dose administration as well as 12 hours away from administration and within two hours of and levels of free copper (using the direct free copper methodologies preciously described herein to measure free copper), total copper, ceruloplasmin and 24S-hydroxy-cholesterol (a copper-implicated oxysterol believed to be produced in the brain that is elevated in the serum of Alzheimer's patients).
  • Results Peak and calculated approximated AUC levels of free copper in the serum of Alzheimer's patients are increased in the immediate release cohort compared to the sustained release cohort, serum levels of total copper and ceruloplasmin are not statistically significant.
  • Example 4 (gastroretentive sustained release zinc): A double blind placebo controlled clinical trial in Alzheimer's patients is designed comparing placebo (Cohort I), 2mg/day immediate release copper supplementation (Cohort II), 100 mg/day gastroretentive sustained release zinc acetate without copper supplementation (Cohort III) and 100 mg/day gastroretentive sustained release zinc acetate with 2 mg/day sustained release copper supplementation (Cohort IV) is carried out for 12 to 24 months.
  • statins such as atorvastatin
  • cholinesterase inhibitors such as donepezil
  • NMDA receptor antagonists such as memantine.
  • the primary endpoint of the study is clinical improvement based upon mini-mental state exam (MMSE) scoring, brain atrophy as measured by volumetric MRI at a minimum of 1.5T resolution using commonly decribed procedures and longitudinal proton magnetic resonance spectroscopy IH-MRS utilizing the PRESS-J and autorepositioning techniques described by Hancu I, et. al (2005).
  • MMSE mini-mental state exam
  • doses of approximately 1 mg/kg/day to 1.5 mg/kg/day bring the levels of available free copper down to negligible amounts.
  • Such doses are then generally titrated in order to maintain available free copper levels (generally measured indirectly based upon systemic levels of ceruloplasmin since copper bound in a tripartite complex with tetrathiomolybdate and albumin are difficulty to distinguish using currently available practical means) within a certain therapeutic range.
  • an object of the present invention is to provide a low but steady concentration of tetrathiomolybdate in the serum to reduce and stabilize free copper levels in the serum through the binding of free originating from copper free copper fluxes(such as those arising from drinking tap water containing soluble copper) solublized copper in tap water), forming a tripartite complex with serum albumin and thus rendering it unavailable to low molecular weight proteins such as homocysteine, capable of crossing through the blood brain barrier.
  • Copper bound to thiomolybdates and albumin are generally too heavy to cross the blood brain barrier, giving copper complexing agents such as tetrathiomolybdate unique pharmacologic advantages as opposed to copper chelators such as d-penicillamine, trientine, cliquinol, EDTA and other metal complexing agents since free copper loosely bound to low molecular weight proteins such as homocysteine easily cross the blood brain barrier creating a burden on the intraneuronal and extraneuronal cuproprotein mechanisms necessary to maintain appropriate levels of copper homeostasis.
  • copper complexing agents such as tetrathiomolybdate unique pharmacologic advantages as opposed to copper chelators such as d-penicillamine, trientine, cliquinol, EDTA and other metal complexing agents since free copper loosely bound to low molecular weight proteins such as homocysteine easily cross the blood brain barrier creating a burden on the intraneuronal and extraneuronal cuproprotein mechanisms necessary to maintain appropriate levels of copper homeost
  • agents such as d-pencillamine and trientine actually serve to also increase the levels of free copper in the CNS due to their nonselective chelation of copper from non-CNS organ compartments such as the liver.
  • thiomolybdates such as tetrathiomolybdate will serve to stabilize and reduce free copper levels in the CNS which have reported to be elevated in Parkisnon's patients as well as downregulate production of the copper and iron binding protein ⁇ -synuclein which comprises the majority of Lewy Bodies associated with such disease as well as other synucleinopathies.
  • the copper binding apoliprotein E which in variant 4 (apoE4) is devoid of two copper binding cysteine residues that limit the ability of apoE to clear copper from the CNS via uptake by the liver.
  • apoE4 copper binding apoliprotein E
  • one skilled in the art could develop other novel formulations of various agents, such as depo injection formulations of copper binding agents bound to various proteins, pegalated peptides containing cysteine residues for example, pegalated SODl, and liposomal complexed copper and iron binding agents capable of accomplishing the same result as the free copper resucing and stabilizing mechanism of action of tetrathiomolybdate and more particularly sustained release tetrathiomolybadte and more particularly low dose sustained release tetrathiomolybdate.
  • an object of the present invention claim the method of using any agent capable of binding and rendering unavailable free copper wherein such agent is made available in the serum at a steady state and preferably does not cross the blood brain barrier.
  • an object of the present invention is to provide stable immediate release pharmaceutical formulations containing thiomolybdates that are useful for chelating and complexing copper in the stomach and gastrointestinal tract and serum.
  • the pharmaceutical formulations and methods described herein are useful to block copper absorption from the intestine while complexing the copper in serum to direct its excretion from the body.
  • compositions for complexing and sequestering the copper in food and liquids, and from endogenous sources, such as saliva, gastric, pancreatic, biliary secretions and sloughed enterocytes, when taken with meals thereby reducing copper absorption as well as having use in complexing and reducing the levels of free copper in serum when taken away from food.
  • endogenous sources such as saliva, gastric, pancreatic, biliary secretions and sloughed enterocytes
  • the present inventions overcomes the conflicting goals of providing an immediate release formulation of thiomolybdates capable of rapidly dispersing in the stomach so as to complex and sequester copper contained in foodstuffs and liquids when taken with meals while at the same time overcoming the inherent instability associated with thiomolybdates so as to provide a pharmaceutical product having a commercially and regulatory acceptable shelf life without compromising the rapid dissolution characteristics necessary to adequately disperse thiomolybdates to complex and sequester copper in the stomach and intestines when taken with meals.
  • the present invention also comprises stable immediate release formulations and methods that when taken away from food are capable of crossing the gastrointestinal digestive tract and chelating and sequestering copper in the serum of a patient in need thereof.
  • Thiomolybdates are comprised of molybdenum and sulfur, and include but are not limited to species such as (MoS 4 ⁇ and (Mo ⁇ 2 S 2 ) 2 -
  • the thiomolybdates can be made as a pharmaceutical acceptable salts, such as, the diammonium salt. These molecules can act as bidentate ligands, and can complex copper. Examples of thiomolybdates include but are not limited to tetrathiomolybdate, trithiomolybdate, dithiomolybdate, and monothiomolybdate.
  • complex thiomolybdates which include but are not limited to a zinc or an iron between two thiomolybdate groups, and which contain thiomolybdate capable of binding or complexing copper.
  • the molecule may have more than four thio groups related to more than one molybdenum.
  • the Group VI transitional metals, tungstate can substitute for molybdate in formulations, as thio- tungstates.
  • a thiomolybdate that has particular relevance for the treatment of neurologically-presenting Wilson's disease is tetrathiomolybdate.
  • Tetrathiomolybdate is a thiomolybdate that comprises a molybdenum atom surrounded by four sulfurs, (MoS 4 ⁇ . Since the 1980's, oral tetrathiomolydate has been the subject of preclinical and human clinical trials conducted by George Brewer, M.D., Fred Askari, M.D. and others for the treatment of initially-presenting neurologic Wilson' s disease as well as other diseases that may benefit from the inhibition of intestinal copper absorption and the sequestration and removal of endogenous copper from the body.
  • Neurologic Wilson's disease is a genetic disease caused by mutations of the ATP7B gene and is characterized by an impaired hepatic ability to incorporate copper into the ceruloplasmin protein and excrete copper yia the bile and stool. This impairment results in elevated levels hepatic and brain copper and that of free copper in the systemic circulation of Wilson's patients which as a result causes toxicities to the liver, brain and other organs.
  • the regimen of tetrathiomolybdate used for initially presenting Wilson' s patients involves up to a sixteen week treatment period in which the patient is sometimes simultaneously administerd Zinc, such as zinc acetate. Treatment can also be followed by daily maintenance therapy with zinc containing agents such as zinc acetate (Galzin®).
  • tetrathiomolybdate is orally administered to patients up to three times daily with food and with additional administration of tetrathiomolybdate given orally away from food.
  • the goal of tetrathiomolybdate administration with food is to act in the stomach and intestine to complex and render unavailable for systemic absorption copper contained in meals, liquids and endogenous copper that enters the gastrointestinal tract.
  • Tetrathiomolybdate is given away from meals to facilitate its systemic absorption thereby allowing it to form a tripartite complex of toxic free copper in serum to serum proteins, such as albumin, and thereby reduce the availability and levels of toxic free copper levels in the serum and its availability to the central nervous system (CNS).
  • CNS central nervous system
  • Reduced availability of copper to the CNS can reduce levels of free copper to normal levels and can prevent or treat psychiatric and neurodenerative toxicities of elevated free copper in the CNS.
  • Te trathiohiomoly dates, under typical atmospheric condition of temperature and humidity are unstable and the active ingredient can undergo oxidation to molybdenum sulfoxide.
  • Molydbdenum sulfoxide is not active as a copper binding agent and therapeutic agent for reduing body copper levels in humans. Tetrathiomolybdates can be expected to lose approximately 10% of its potency within three months if stored under such conditions. Accomplishing the dual goal of creating an oral thiomolydate pharmaceutical product capable of immediately releasing and rapidly dispersing in the stomach to complex copper contained in meals and liquids while at the same time creating a pharmaceutical product having acceptable stability, shelf life and consistent potency has been the subject of considerable efforts by others in the field. For example, see: U.S. Patent Application Ser. No. 11/256,540, filed October 21, 2005 by Ternansky et.
  • the current invention in one aspect comprises immediate release capsules containing a thiomolydate wherein the capsule has a very low water content, wherein the capsule contains an exipient that has a low water content and wherein the capsules are packaged in containers or blister packs, to protect the thiomolybdate from the atmospheric oxygen by purging and sealing the capsules with an inert gas, such as nitrogen or argon.
  • an inert gas such as nitrogen or argon.
  • the packaging of the capsules may contain one or more chemical indicators to alert the use of a breach of the anhydrous or anaerobic seal of the package with an indicator, such as one for moisture or oxygen.
  • the present invention also provides a method of timing the administration of a thiomolybdate immediately prior to eating and co-administration of an adequate volume of a liquid to assure the rapid dissolution and the broadest complexation coverage of copper containing foods and liquids.
  • the present invention also includes the use of low dose sustained release tetrathiomolydate or any other comparable agent as so described to treat other non- CNS disorders in which oxidation due to elevated peak and AUC free copper are known to play a role, such as atherosclerosis and liver diseases such as NASH, non- viral hepatitis and diabetes.
  • sustained release formulations may preferentially comprise two or more microparticle or matrix types contained in a single pill capsule or tablet for example.
  • Such pill or capsule may contain an immediate release form, while one enteric coated particle is designed to release in pH environment of the jejunum and small intestine for example and yet another designed to release in environment of the ileum and colon.
  • the present invention also comprises a pharmaceutical package whereby each pill, tablet or capsule is separately sealed in its own modified atmosphere packaging such as an impervious foil pouch or cold form blister pack that is purged under argon or nitrogen gas to expel 98% or greater of atmospheric air and moisture until ready for use.
  • capsules containing tetrathiomolybdate should also be specifically selected for low moisture content so to limit interaction between the capsule and active ingredient.
  • Pouches or blister pack packaging can be further packaged in an inert purged package together with an desiccant and sacrificial oxidant to improve stability.
  • the size of the tetrathiomolybdate crystals are controlled to provide selected, sustained or delayed release in the stomach and the intestines. Crystal size may be controlled through controlled crystal growth, or by milling as described below.
  • tetrathiomolybdate crystals for example, ammonia tetrathiomolybdate pass through the stomach essentially unchanged and dissolve in the gut, while smaller crystals dissolve in the stomach.
  • sustained release in relation can be achieved by providing a mixture of crystals of varying sizes.
  • a mixture comprising crystals of 50-100 microns and 200-500 microns may be provided, in a single capsule.
  • ATTM ammonium tetrathiomolybdate
  • ATTM purchased from Sigma- Aldrich was sieved using a 1700 micron screen. Crystals which did not pass through the screen were called large and used in the experiment. 2. ATTM crystals were ground so that no crystals were larger than 50 microns. These will be referred to as small crystals. 3. 100 mg of each type of crystal was weighed and added to separate 1.7 ml plastic centrifuge tubes. 4. 1 ml of buffer pH 2 was added to each tube without mixing. 5.
  • each tube was centrifuged at 14,000 rpms for 1 minute. 6. The supernatant was removed and analyzed for ATTM content. 7. To the pellet of each tube was added 1 ml of phosphate buffer pH 7.4 8. Each solution was vortex-mixed. 9. Each tube was centrifuged at 14,000 rpms for 1 min. 10. The supernatant from each tube was analyzed for ATTM content. Analysis: 1. A 1 to 100 dilution of all supernatants were made in phosphate buffer pH 7.4. 2. 2 ml of each diluted sample were added to plastic cuvettes. 3. Absorbance was measured at 467 nm. 4. The extinction coefficients (EC) of ATTM for each sample were calculated. Results:
  • the small crystals are placed in the same type of solution and immediately release a strong odor of sulfur and form a disaggregate cloudy black solution.
  • the dissolution properties of the large verses small crystals are clearly distinct.
  • a methylcellulose capsule containing 20 mg of tetrathiomolybdate and 180 mg of excipient containing a heterogenous size range ( 200 -800 microns) of tetrathiomolybdate crystals was examined in a dissolution chamber initially containing simulated stomach acid. The gelatin capsule dissolved within 15 minutes. Tetrathiothiomolybdate recovery was assessed by UV/Visible absorption periodically over 3 hours, a time consistent with the initiation of gastric emptying and the initial phase of postprandial absorption.
  • Tetrathiomolybdate was not detectable in the simulated gastric samples removed and neutralized at any time point over this three hour period, suggesting the agent did not dissolve or was disaggregated.
  • the remaining gastric juice was made alkaline by adjusting the pH to 8-9, the solution turned orange to red and was assessed by UV/Vis spectroscopy to determine the recover of tetrathiothiomolybdate, which was approximately 1 % of the amount in the capsule.
  • Tetrathiothiomolybdate crystals are either crushed to a powder, that is initially orange in color, or placed in water to dissolve yielding a bright orange solution, which is filtered and lyophilized, which results in a dry powder orange in color.
  • the crushed crystals are purged with nitrogen overlay and slowly turn black within a few day at room temperature in a closed tube, while the lyophilized dry powder under similar conditions remains orange for over a month. This experiment suggests the thorough removal of water from tetrathiothiomolybdate improves stability to the atmosphere.
  • Example 7 Similar amounts of lyophilized tetratiomolydate powder is dissolved in water, placed in a capsule, prepared as a non-coated tablet, or enteric coated tablet and dosed once a day in fed or fasted rats for one week. At the end of the multiple dosing period, blood samples are taken periodically over 24 hours for pharmacodynamic determination of ceruloplasmin and pharmacokinetic determination of molybdenumum.
  • the primary endpoint of the study is clinical improvement based upon cognitive assessments such as mini-mental state exam (MMSE) scoring, brain atrophy as measured by volumetric MRI at a minimum of 1.5T resolution utilizing commonly described procedures and longitudinal proton magnetic resonance spectroscopy IH- MRS utilizing the PRESS-J and autorepositioning techniques described by Hancu I, et. al (2005).
  • MMSE mini-mental state exam
  • IH- MRS longitudinal proton magnetic resonance spectroscopy
  • MMSE mini- mental state exam
  • Cohort II shows less decline in all parameters as compared to Cohort I and potential objective indications improved metabolic brain function in as little as 3-6 months as compared to Cohort I.
  • Molybdenum and sulfur containing formulations An object of the present invention includes oral nutriceutical formulations that contain elemental molybednum and sulphur. Upon administration and dissolution in the stomach such formulations would combine in the low pH environment of the stomach and form thiomolybdates capable of complexing free copper in the gastrointestinal tract and systemic circulation much like ammonium tetrathiomolybdate. While such formulations would be expected to be far less reliable in terms of dosing, they might have the advantages as natural products purusnat to the Dietary Health Supplement and Education Act (DSHEA).
  • DSHEA Dietary Health Supplement and Education Act
  • a sustained release trace metal of the invention may be incorporated into sustained release pharmaceutically acceptable sustained release microsphere, matrix, pellet or particle (all of which are commonly known in the art) in the form as a pure cation or salt
  • a pharmaceutically acceptable, stable, natural or synthetic carriers to which such metals are known to bind, such as, for example, plant fiber, whey, metallotheionein, transferrin, proteins and/or milk or milk by- prodcuts.
  • Such carriers will have the benefit of further inducing the gradual digestion and absorption of the trace metals as they are naturally found in foods.
  • the invention thus further provides a gastroretentive and/or sustained release pharmaceutical formulation incorporating one or more trace metals, such as zinc, copper and iron together with one or more pharmaceutically acceptable carriers and, optionally, other therapeutic and/or prophylactic ingredients.
  • the carrier(s) must be "acceptable” in the sense of being compatible with the other ingredients of the formulation and not deleterious to the recipient thereof.
  • Another object of the present invention is to provide a copper supplement that may be administered in conjunction with, or combined with, a copper malabsorption agent, wherein the copper is in a form bound to or formulated with lipids, whey or casein.
  • a copper supplement formulated in such a fashion would be intended to mimic, in pill, capsule or liquid form, the manner in which copper is provided from mother to infant via human breast milk.
  • Such copper upon reaching the stomach and small intestines, would be appropriately processed in a proper digestive manner, as opposed to copper which is in water or in a pure salt form. It is anticipated that the processing of such a copper supplement would mimic the high bioavailability found in breast milk (24%) or cow milk (18%), while also permitting the normal digestion and processing of copper by the intestines and liver, thereby reducing the level of burden of free or loosely bound copper in the serum and CNS.
  • Such formulations could also include other essential metals and minerals such as iron or zinc.
  • a copper and/or iron supplement formulated with lipids, whey and other proteins, with which copper is normally found in breast milk may be formulated with a copper malabsorption agent such as zinc so as to simultaneously provide a bioavailable amount of copper and/or iron in a form for normal processing by the intestines, while at the same time inducing the production of metallothionein in the intestines to block and protect against the subsequent absorption of ionic copper from drinking water.
  • such carrier bound copper supplement is incorporated within a sustained release microparticle or matrix so as to further regulate the absorption, and reduce the potential to cause peak elevated levels of free copper in the systemic circulation and CSF.
  • Such formulations should reduce or avoid the need to monitor patients for hypocupremia or anemia, and also lower the levels of free or loosely bound serum or CNS copper while bolstering the levels of ceruloplasmin bound copper (given its processing by the liver by virtue of the first pass effect and normal copper handling, which mimics that of the evolutionarily proven copper and metal supplementation methods by which a mother processes and passes nutritional copper and other metals to a newborn baby via breast milk).
  • Such formulation could also include other essential trace elements in a carrier bound complexed sustained release formulation, such as, iron, calcium, molybdenum, selenium, and magnesium, for example.
  • Complexed sustained release calcium formulations are useful for the treatment or prevention of diseases involving calcification such as in the arteries, kidneys, lungs and brain.
  • stable copper isotopes Cu 65 and Cu 67 are utilized as the active copper ingredient to permit copper balance and treatment effect to be evaluated and adjusted on an individual basis.
  • Such studies may be conducted by sampling serum, CSF fluid, stool, or urine, or by biopsy.
  • Such information provides a useful diagnostic method to evaluate the effects of other therapeutic approaches, such as therapies to improve hepatic excretion of copper as well as copper chelators.
  • the zinc and copper formulations described herein may be administered as a nutritional supplement, parenterally, by immediate release, or by depo release injection.
  • Oral nutritional formulations of zinc and complexed copper may also contain certain copper absorption enhancing agents such as glycerol and NaCl, or gum arabic, to increase the bioavailability of complexed copper.
  • the copper and zinc formulations described herein may also be useful for the treatment of gastrointestinal disorders such as chronic diarrhea, diarrhea predominant irritable bowel syndrome and infections for which zinc is known to be beneficial.
  • the present invention may be administered as a two-pill system whereby complexed copper pills or formulations are orally administered first and are followed by orally administered zinc-containing pills either together or after sufficient delay.
  • the present invention also contemplates incorporation of other essential minerals for which intestinal zinc may also reduce bioavailability, such as iron and cadmium.
  • the present invention may be preferentially combined with a cholesterol lowering agent for the treatment or prevention of atherosclerosis, dementia, Alzheimer's disease and other neurogenerative diseases, including neuromuscular diseases, which are associated with abnormal accumulation of copper associated proteins, such as juvenile and sporadic inclusion body myositis and myositis of the elderly.
  • a further embodiment of the present invention contemplates the use of the zinc formulations described herein or copper lowering agents for the treatment of neurologic and psychiatric manifestations of hepatic diseases associated with impaired liver copper excretion, such as colangitis, hepatitis and cirrhosis, for example, in which free or loosely bound serum or CSF copper is elevated.
  • Example 10 Example 10.
  • a pill or capsule containing high-density 100 mg zinc salt pellets capable of retention in the walls of the stomach is administered to groups of individuals in the morning.
  • a control group of individuals receives a 100 mg capsule of normal zinc acetate, while another control group receives a placebo.
  • Such individuals consume copper-containing or Cu 64 containing distilled or tap water at identical times throughout the day.
  • Serum samples are obtained before, during, and after each drinking event. Such samples are immediately measured for their respective free or loosely bound copper content utilizing total serum copper less copper bound to ceruloplasmin, direct measurement copper bound to various known and unknown proteins and peptides by means of separation based upon molecular weight, as per the column method described by Bohrer D (2004) or MALDI-TOF described by Sarkar E (2004).
  • the placebo group shows the highest peak levels of free or loosely bound copper
  • the bolus oral zinc group has a lower peak free or loosely bound copper level
  • the gastroretentive/sustained release zinc group shows the lowest levels of free or loosely bound free copper.
  • Example 11 Copper, copper salts or copper bound to low molecular weight amino acids is dissolved in a solution of dried or evaporated milk, dried whey, dried milk lipids, or dried milk proteins or other natural copper binding proteins. The resulting mixture of copper bound complexes may be dried and formulated as a pill or tablet.
  • the dried copper bound complexes are formulated in a pill or capsule together with zinc.
  • the pill or tablet is formulated utilizing gastrorententive forms of zinc, enteric coated zinc and/or sustained release zinc such that the copper bound complexes are released into the gastrointestinal tract ahead of the zinc.
  • the copper bound complexes are digested in the GI and absorbed by enterocytes where they are processed intracellularly with metallothionein and/or transcuprein, whereupon such metallothionein and/or transcuprien bound copper enters the liver and is taken up for processing and incorporation into ceruloplasmin and then either released into the serum, retained in the liver or excreted into the bile.
  • the later passing zinc is absorbed in the enterocytes, thereby upregulating metallothionein, which serves to block subsequent absorption of solubilized copper (free or loosely bound copper such as copper-containing tap water or other liquids that would otherwise pass into the hepatic circulation upon water flux without proper processing by the enterocyte and enter the hepatic circulation bound to albumin).
  • solubilized copper free or loosely bound copper such as copper-containing tap water or other liquids that would otherwise pass into the hepatic circulation upon water flux without proper processing by the enterocyte and enter the hepatic circulation bound to albumin.
  • a cross-over design is utilized, whereby naturally occurring copper (39% Cu 63 and 69% Cu 65 ) is administered orally as a pill as described above or solubilized in water. Such administration may be short, or for up to 90 or more days in the case of the stable copper isotopes.
  • Portal and/or serum samples are collected and fractionated into components utilizing columns capable of separating ceruloplasmin, transcuprien, albumin, small proteins and peptides.
  • the copper isotopes bound to each component are measured, demonstrating a statistically significant greater proportion of ceruloplasmin and transcuprien bound copper isotopes administered in the protein bound form as compared to the solubilized form, while the copper isotopes administered solubilized in water show a greater percentage loosely bound to albumin and other low kinetic binding proteins and amino acids.
  • This effect is ameliorated with the pill or capsule formulation which contains early release copper and gastroretentive zinc, as the zinc protects against subsequent absorption of free copper solubilized in water, while reducing the tendency for hypocupremia.
  • Example 13 Dissolution of ATTM as a Capsulated Formulation Introduction: We performed a dissolution experiment using a capsule containing 20 mg of ammonium tetrathiomolybdate plus excipient. ATTM crystals used were 500 microns or smaller. No ATTM was recovered in the gastric dissolution medium at any time.
  • Dissolution time 3 hours Procedure 1. A #2 QualiCaps capsule available from Shionogi Europe, B. V. was mechanically dropped into media at time zero 2. 1 ml of media was then collected at 1, 2, 4, 8, 16, 30, 45, 75, 120 and 180 min 3. Following 3 hours of dissolution, the pH was raised to 9 using NaOH pellets. 4. The media was paddled for 1 hour. 5. After 1 hour a sample was collect for analysis Analysis 1. 400 ul of collected media was diluted to 2 ml with phosphate buffer, pH 7 2. The solution was placed in a cuvette and scanned between 260 and 600 nm. 3. Absorbance was measured at 367 nm. 4. The extinction coefficient was calculated using Beer's law. 5.
  • the extinction coefficient of a reference sample in a capsule is 13500 cm "1 M "1 Conclusion: 1. Any ATTM that dissolves in media at pH 2 is immediately destroyed. 2. Some of the ATTM floated in the media as black specks following 3 hours of dissolution. 3. After three hours when the pH of the solution was raised to pH 9, the color of the solution changed to an orange color. 4. This presumably is stable ATTM that has dissolved. 5. The amount of ATTM remaining was only about 10% of theory. 6. Therefore 90% of the original dose of ATTM was destroyed in the artificial gastric fluid over three hours. 7. With enteric coating, much lower doses may be required to achieve the same therapeutic effect.
  • ATTM was 60% to 90% stable after 8-Weeks when stored at 40 oC and 70% RH.
  • the worst formulation was gel caps, room air, and lactose anhydrous.
  • ATTM formulations were tested at room temperature and ambient humidity. Conditions were Mono (monohydrate lactose), Anydrous (anhydrous lactose, Quali (methyl cellulose capsules), Air (air atmosphere) and N2 (nitrogen atmosphere.
  • Various procedures are possible for both preparing the ammonium tetrathiomolybdate (ATTM) or ammonium polysacharide (API), preparing formulations and capsules containing same, and the use of same including: Milling ammonuim tetrathiomolybdate (ATTM) L A method for milling ATTM under condition of low moisture and oxygen levels that separates ATTM from oxidized forms of the active ingredient that present as black tar- like by-products (flaky tar). 2.
  • Excipient Selection 14 A method of formulation whereby the ammonium polys acharide (API) is mixed with an excipient that enhances the pharmaceutical properties of the API. 15.
  • An example of 15 but not limited to the example is anhydrous lactose.
  • An example of 17 but not limited to the example is a crown ether. 19.
  • Salt Selection 21. A method of formulation whereby the API forms a salt with a cation that enhances or protects the pharmaceutical properties of the API. 22.
  • An example of 22 but not limited to the example is the nicotinic ester of tocopherol.
  • 24. A formulation of 21, in which the cation is hydrophobic and acts to prevent the hydration of the API. 25.
  • An example of 24 but not limited to the example is benzylamine. Agents added to the Excipient which Prevent the Acid Hydrolysis of the API 26.
  • 27. A formulation of 26 in which stomach acid hydrolysis of API is prevented with the addition of an anti-acid to the excipient.
  • 28. A formulation of 26 whereby an anti-acid is administered slightly before, simultaneously, or slightly after, but separately from the ATTM pharmaceutical formulation. 29.
  • An example of 27 but not limited to the example is sodium bicarbonate.
  • An example of 28 but not limited to the example is sodium bicarbonate.
  • An example of 32 but not limited to the example is an aminopolysacharide (API).
  • An example of an aminopolysacharide is polyglucoseamine or chitosan.
  • 36. A method of 35 where some or all of the API is enteric coated as microtablets that are encapsulated in a capsule. 37.
  • 37. A method of 35 where the enteric coating is variable to afford variable rates of dissolution of the microtablets.
  • 38. A method of 35 where a single tablet is enteric coated for dosing.
  • 39. A method of 35 whereby tablets of various enteric coatings are prepared for administration to a patient such that the enteric coatings release the API at different rates. 40.
  • 41. A method of 40 where a copper lowering mediation is but not limited to a zinc acetate, zinc sulfate, tetrathiomolybdate, tetrathiotungstate, penecillamine, or trientine. Agents added to the Excipient which Aid in the absorption of the API.
  • 42. A method of formulation whereby an agent as part of the excipient is added to the API so that a complex is formed which provides for enhancing absorption of the API.
  • 43. An example of 42 but not limited to the example is an aminopolysacharide.
  • An example of an aminopolysacharide is polyglucoseamine or chitosan. 44. A mechanism of 43 in which chitosan added to the excipient forms an insoluble colloidal complex with the API following dissolution in stomach acid and is metabolically converted into di- or tri-saccharides complexes with API that suitable substrates for absorption.
  • 53. A method of coating in 51 whereby API resists dissolution in the stomach but readily dissolves in the upper and lower intestines. Combined Targeting of Specific Organs for API Dissolution 54.
  • 55. A formulation of 54 which utilizes a combination of release mechanisms.
  • 56. An example of 55 in which API release is targeted for dissolution in the stomach. Dissolution in the stomach is desired for complexation of copper in the stomach from food and drink as a preventative measure in terms of copper absorption occurring later in the intestines.
  • An example of 56 would be finely powdered API and excipient contained in a capsule that readily dissolves in stomach acid. The excipient may contain an API complexing agent which prevents acid hydrolysis but allows for API interaction with free copper in the stomach. 58.
  • An example of 61 but not limited is sodium alginate.
  • Sodium alginate in powder form is a common food stuff that is known to bind heavy metals and prevent their systemic absorption.
  • a mechanism of 61 would involve the use of sodium alginate to bind free copper from food in the stomach while allowing for coated API to dissolve in the intestine for systemic absorption.
  • Formulation of API that Provides for Sustained Release 64 A method of formulation which allows for sustained release of API in the stomach.
  • 65. A formulation of 64 in which API is mixed with an agent that is semi- resistant to dissolution in stomach acid while also possessing an overall material density that provide for buoyancy in stomach fluids.
  • a non- limited example of 65 would be the formulation of API in a digestible wax.
  • the density of the wax would provide for buoyancy in stomach fluids and the slow digestibility of the wax would provide for the sustained release of API.
  • Functional Features 67. A method to reduce the number of times a copper reducing therapy is orally administered to a patient. 68. A method in which ATTM is administered to a patient in which the absorption of the API is increased. 69. A method in which a formulation of ATTM is administered to a patient to enhance the pharmacodynamic profile of the agent at an equivalent dose. 70. A method in which a formulation of ATTM achieves indications of serum copper, serum ceruloplasmin, urinary copper, or hepatic copper lowering at a lower dose. 71. A method in which a formulation of ATTM achieves an indication of copper more rapidly than an equivalent dose.
  • Monitoring and Dosing 72 A method by which a patient or caregiver monitors the effectiveness of an ATTM containing formulation or dose program by assaying a body fluid, excretion or organ for a therapeutic marker indicative of effective copper reduction therapy.
  • 72 A method of 72 where the body fluid is urine.
  • 72 A method of 72 where the body fluid is blood or blood serum.
  • 75 A method of 72 where the body fluid is blood plasma.
  • 76 A method of 72 where the body fluid is saliva.
  • a method of 72 where the body fluid is cerebral spinal fluid.
  • 78. A method of 72 where the material assayed is stool. 79.
  • a method of 72 where the material assayed is liver. 80.
  • longitudinal cognitive assessments of a patient are made to indicate a slowing of progression or regression of disease.
  • quality of life assessments of a patient are made to indicate a slowing of progression or regression of a disease.
  • Diseases for treatment 83.
  • the central nervous disease can be, but is not limited to; Wilson's disease, Alzheimer's disease, Huntington's disease, Schizophrenia, Parkison's Disease, ALS, and Prion diseases.
  • the inflammatory disease can be, but is not limited to; psorisis, rheumatoid arthritis, lupus, inflammatory bowel disease, etc. 87.
  • a method of 84 where the fibrtotic disease can be, but is not limited to; idiopathic pulmonary fibrosis, primary biliary cirrhosis, nonalcoholic steatohepatits, liver cirrhosis, glomerulonephritis, systemic fibrosis and rheumatoid arthritis, post-surgical adhesions, adult respiratory distress syndrome (ARDS); coal workers' pneumoconiosis (CWP); Hermansky- Pudlak syndrome (HPS); systemic sclerosis (SS), tumour stroma in lung cancer, and obliterative bronchiolitis (OB) after lung transplantation.
  • ARDS adult respiratory distress syndrome
  • CWP coal workers' pneumoconiosis
  • HPS Hermansky- Pudlak syndrome
  • SS tumour stroma in lung cancer
  • OB obliterative bronchiolitis
  • 90. A method of formulation in which protein is added as part of the excipient.
  • a method of 54 in which the stable API-protein complex is capable of complexing copper in the stomach. 93.
  • An example of a formuation of 54 would be that bovine serum albumin is added to the excipient.
  • a mechanism of 54 would be that the API and the bovine serum albumin would form a stable complex capable of complexing with copper in the stomach.
  • An example of a fo ⁇ nuation of 54 would be a synthetic D-polypeptide which complexes with copper in which the complex can further complex with copper and exit the GI tract intact and is not digested. 98.

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Abstract

L'invention concerne des compositions et des méthodes dans lesquelles sont utilisés des moyens permettant d'obtenir et de maintenir un taux de cuivre cible chez des personnes, afin de traiter et de prévenir des maladies liées au taux de cuivre sérique.
PCT/US2007/060345 2006-01-10 2007-01-10 Compositions pharmaceutiques et méthodes permettant d'obtenir et de maintenir un taux de cuivre cible, stable, et de prévenir et de traiter des maladies du système nerveux central liées au taux de cuivre sérique WO2007084818A2 (fr)

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AU2007205996A AU2007205996A1 (en) 2006-01-10 2007-01-10 Pharmaceutical compositions and methods to achieve and maintain a targeted and stable copper status and prevent and treat copper-related central nervous system diseases
CA002675230A CA2675230A1 (fr) 2006-01-10 2007-01-10 Compositions pharmaceutiques et methodes permettant d'obtenir et de maintenir un taux de cuivre cible, stable, et de prevenir et de traiter des maladies du systeme nerveux centralliees au taux de cuivre serique
EP07718298A EP1993607A4 (fr) 2006-01-10 2007-01-10 Compositions pharmaceutiques et méthodes permettant d'obtenir et de maintenir un taux de cuivre cible, stable, et de prévenir et de traiter des maladies du système nerveux central liées au taux de cuivre sérique
JP2008550493A JP2009523175A (ja) 2006-01-10 2007-01-10 目標とされる安定な銅状態を達成及び維持して銅関連中枢神経系疾患を予防及び治療するための医薬組成物及び方法

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US20070207191A1 (en) 2007-09-06
AU2007205996A1 (en) 2007-07-26
WO2007084818A3 (fr) 2009-01-08
EP1993607A2 (fr) 2008-11-26
JP2009523175A (ja) 2009-06-18

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